Regulation of rpoS gene expression in Pseudomonas: involvement of a TetR family regulator

The rpoS gene encodes the sigma factor which was identified in several gram -negative bacteria as a central regulator during stationary phase. rpoS gen e regulation is known to respond to cell density, showing higher expression in stationary phase. For Pseudomonas aeruginosa, it has been demonstrated that the cell-density dependent regulation response known as quorum sensing interacts with this regulatory response. Using the rpoS promoter of P. put ida, we identified a genomic Tn5 insertion mutant of P. putida which showed a 90% decrease in rpoS promoter activity, resulting in less RpoS being pre sent in a cell at stationary phase. Molecular analysis revealed that this m utant carried a Tn5 insertion in a gene, designated psrA (Pseudomonas sigma regulator), which codes for a protein (PsrA) of 26.3 kDa. PsrA contains a helix-turn-helix motif typical of DNA binding proteins and belongs to the T etR family of bacterial regulators. The homolog of the psrA gene was identi fied in P. aeruginosa; the protein showed 90% identity to PsrA of P. putida . A psrA::Tn5 insertion mutant of P, aeruginosa was constructed, In both Ps eudomonas species, psrA was genetically linked to the SOS lexA repressor ge ne. Similar to what was observed for P. putida, a psrA null mutant of P. ae ruginosa also showed a 90% reduction in rpoS promoter activity; both mutant s could be complemented for rpoS promoter activity when the psrA gene was p rovided in trans, psrA mutants of both Pseudomonas species lost the ability to induce rpoS expression at stationary phase, but they retained the abili ty to produce quorum-sensing autoinducer molecules. PsrA was demonstrated t o negatively regulate psrA gene expression in Pseudomonas and in Escherichi a coil as well as to be capable of activating the rpoS promoter in E. call. Our data suggest that PsrA is an important regulatory protein of Pseudomon as spp. involved in the regulatory cascade controlling rpoS gene regulation in response to tell density.